Oxygen is a critical element to many chemical and physiological processes. Methods and apparatuses for measuring the presence and/or concentration of oxygen is important to many fields of endeavor. For example, microorganisms consume oxygen during growth, depleting the concentration of oxygen in the surrounding environment. Oxygen tension in mammalian cell culture can profoundly affect cellular differentiation, viability, and proliferation. Therefore, a measurement of oxygen in the environment surrounding a cell culture provides data on the cells in the culture.
Oxygen has been monitored with electrochemical sensors. Electrochemical sensors may allow continuous monitoring of oxygen. For example, electrochemical methods for detection of oxygen have been used in bio-reactors to insure sufficient media oxygenation. However, it may be substantially impossible to place the oxygen measurement electrode near the cells in culture to know the local conditions since the electrochemical sensors are typically too large. Electrochemical sensors are typically used in production-scale applications, and routine observation of the cells in culture is difficult if not impossible. Even when desired, electrochemical detection is not easily compatible with common microscopic integration of the cells. Also, the electrochemical approach is difficult to parallelized for high-throughput applications. Additionally, electrochemical sensors may be prone to chemical and electrical interference; they may consume oxygen; and typically have a large bulk mass. Therefore, in many applications, electrochemical sensors may be an unsatisfactory method of oxygen measurement.
Optical sensors for oxygen have overcome some of these problems associated with electrochemical sensors. Optical sensors may be based on a change in luminescence intensity emanating from phosphorescent compounds or fluorophores which are quenched with oxygen. Commercially available phosphorescent oxygen sensors typically utilize ruthenium based fluorophore to measure changes of oxygen. However, these fluorophores may lack the sensitivity to measure small changes in oxygen level due to the inherently low quantum yields. In addition, many of these fluorophores may be unstable, exhibit rapid photobleaching, and/or may load unevenly across culture dishes. Additionally, dye dispersed in aggregates may significantly compromise the ability to integrate alternative microscopy strategies such as phase contrast imaging and fluorescence imaging.
Most recently, platinum based fluorophores have been used in optical sensors to overcome some of the problems associated with the ruthenium based fluorophores. However, platinum based fluorophores may exhibit a cytotoxic effect on the growth of the cells in the culture media. The cytotoxicity of the platinum based fluorophores may thus cause the phosphorescent oxygen sensor to fail to provide desired representative data of the cell culture.
Precise measurement of dissolved oxygen in a cell culture environment in real time remains difficult. What is needed is an apparatus and method that overcomes some of the aforementioned problems associated with the prior art.